Thymic epithelial cells (TECs) comprise a diverse group of cells that form a complex microenvironmental network required for T cell development and repertoire selection. The Program Project, Project 2 focuses on pathways and mechanisms involved in TEC development, homeostasis and involution, in this case with particular emphasis on medullary epithelium. Development of a functional medullary epithelial compartment is vital for establishing central tolerance. Medullary TECs (mTECs) are unique in their expression of a highly diverse array of tissue restricted antigens (TRAs) that are required to negatively select self-reactive thymocytes. Widespread autoimmune disease occurs in patients or mice that inherit genetic defects in TRA expression. In addition, our lab and others have shown that autoimmunity develops in mice with genetic defects that prevent mTEC development during ontogeny. The incidence of autoimmunity increases with age. Therefore, an attractive hypothesis is that progressive loss of a previously established mTEC compartment during thymus involution compromises central tolerance, increasing the risk of developing autoimmune disease with advancing age. However, the precise mechanisms that regulate their development and homeostasis, as well as their lineage relationships and functional potential remain to be determined. A strategy to prevent or reverse loss of the mTEC population with age also requires identification of the molecular pathways that regulate mTEC differentiation and proliferation. We also find that expression of a constitutively activated StatS transgene regulated by a K5 promoter induces mTEC development in RAG-2-/- mice that normally are devoid of organized medullary regions due to an early block in thymocyte differentiation. This suggests that manipulating StatS activation in mTEC precursors could be a viable approach to sustain or restore the mTEC compartment during aging. To investigate the regulation of mTEC postnatal homeostasis and function, Project 2 will establish lineage relationships among mTEC subsets, identify the consequences imposed by age-associated loss of the mTEC compartment, and establish molecular pathways that sustain or promote mTEC development. RELEVANCE (See instructions): Age-related thymus involution is associated with a decline in mmune function and increased autoimmunity. We propose that loss of mTECs during thymys involution impairs negative selection of self-reactive thymocytes leading to autoimmunity. We will elucidate molecular mechanisms regulating mTEC homeostasis to identify targets for therapeutic intervention aimed at preventing thymus involution and mTEC loss.